METHODS FOR IDENTIFYING AGENTS THAT MODULATE p44

Abstract

The present invention provides methods for identifying evolutionarily significant polynucleotide and polypeptide sequences in human and/or non-human primates which may be associated with a physiological condition, such as enhanced resistance to HCV infection. The invention also provides methods for identifying evolutionarily significant polynucleotides with mutations that are correlated with susceptibility to diseases, such as BRCA1 exon 11. The methods employ comparison of human and non-human primate sequences using statistical methods. Sequences thus identified may be useful as host therapeutic targets and/or in screening assays.

1 . A method for identifying an agent as a candidate for increasing human p44 function, comprising:
 contacting at least one candidate agent with a p44 polypeptide selected from the group consisting of   (a) a human p44 polypeptide comprising SEQ ID NO:36;   (b) a polypeptide encoded by a polynucleotide comprising SEQ ID NO:34;   (c) a polypeptide encoded by a polynucleotide consisting essentially of nucleotides 1-457 of SEQ ID NO:34, and   (d) a polypeptide consisting essentially of amino acids 1 to 152 of SEQ ID NO:36; and   detecting whether the agent binds to the human p4-4 polypeptide encompassing least one of the following positions: Arg36, Ser68, Glu71, Gly72, Asp84, Cys95, and Thr106.   
     
     
         2 . A method for identifying an agent that increases a human or chimpanzee p44 function, comprising:
 contacting at least one candidate agent with a p44 polypeptide selected from the group consisting of:
 (a) a polypeptide encoded by a polynucleotide comprising nucleotides 1-457 of SEQ ID NO:34 or SEQ ID NO:31, and; 
 (b) a polypeptide having at least one amino acid change at positions 36, 68, 71, 72, 84, 95, and 106, but is otherwise identical to amino acids 1 to 152 of SEQ ID NO:36 or SEQ ID NO: 34; and 
   (ii) detecting a p44 polypeptide function,
 wherein said agent is identified by its ability to increase the human or chimpanzee p44 polypeptide function relative to the function of the p44 polypeptide in the absence of the candidate agent. 
   
     
     
         3 . The method of  claim 2 , wherein the p44 function is selected from the group consisting of microtubule assembly and resistance to HCV infection. 
     
     
         4 . The method of  claim 2  wherein the p44 polypeptide function to be increased is human. 
     
     
         5 . The method of  claim 4  wherein the increase of the p44 function results in a human function that is more similar to a chimpanzee p44 function. 
     
     
         6 . The method of  claim 4  wherein said candidate agent is a small molecule that forms a complex with human p44 that mimics the three-dimensional structure of the chimpanzee p44, whereby the human p44 function is increased. 
     
     
         7 . The method of  claim 4  wherein said candidate agent is a small molecule that interacts with human p44 amino acids so as to increase the human p44 polypeptide function. 
     
     
         8 . The method of  claim 1 , wherein said candidate agent is a small molecule that binds with at least one human p44 amino acid selected from the group consisting of Arg36, Ser68, Glu71, Gly72, Asp84, Cys95, and Thr106 of SEQ ID NO:36. 
     
     
         9 . A method for identifying an agent that decreases susceptibility to a human's hepatitis C virus (HCV) infection, comprising:
 (i) contacting at least one candidate agent with a p44 polypeptide capable of decreasing susceptibility to a human's HCV infection selected from the group consisting of:
 (a) a polypeptide encoded by a polynucleotide comprising nucleotides 1-457 of SEQ ID NO:34 or SEQ ID NO:31, and 
 (b) a polypeptide having at least one amino acid change at positions 36, 68, 71, 72, 84, 95, and 106, but is otherwise identical to amino acids 1 to 152 of SEQ ID NO:36 or SEQ ID NO: 34; and 
   (ii) detecting susceptibility to HCV infection in in vitro hepatocytes or in vivo animal model.   
     
     
         10 . The method of  claim 9 , wherein said agent is identified by its ability to decrease susceptibility to HCV infection relative to the susceptibility to HCV infection in the absence of the candidate agent. 
     
     
         11 . The method of  claim 3 , wherein increased microtubule assembly is detected via antibody detection of enhanced p44 assembly into microtubules in cultured hepatocytes. 
     
     
         12 . The method of  claim 3 , wherein increased resistance to HCV infection is detected via decreased viral titer in in vitro hepatocytes or an in vivo animal model.